The inflammatory milieu is a decisive factor in autoimmunity and yet we know very little about how the tissue environment contributes to immune regulation. This knowledge is essential if we are to devise strategies that effectively suppress autoimmunity in inflamed islets and prevent type 1 diabetes (T1D). We have recently determined that high molecular weight hyaluronan (HMW-HA), a component of the extracellular matrix (ECM) of healing tissues, promotes the stability and function of FoxP3+ regulatory T-cells (Treg). HMW-HA does this by crosslinking CD44 and substituting for IL-2 in the IL-2R/STAT5 signaling Treg required for Foxp3 expression and production of IL-10, a key immunoregulatory cytokine. Low molecular weight hyaluronan (LMW-HA), generated from HMW-HA catabolism during infection and chronic inflammation, cannot crosslink CD44 and inhibits Treg function. These data support a model whereby HA integrity governs Treg function in injured and healing tissues. This model predicts that receptivity to HMW-HA signals may govern regulatory T-cell function in inflamed tissues. Indeed, we find that FoxP3+ Treg from T1D subjects have diminished expression of CD44v6, a CD44 variant isoform involved in HMW-HA binding. Reduced CD44v6 expression might impair receptivity to HMW- HA tissue integrity signals and thereby undermine Treg function and persistence in vivo. Our model also predicts that it may be possible to prevent autoimmunity by supporting HA integrity. In healing tissues, HMW-HA degradation is prevented by TSG-6, an HA-binding molecule (hyaladherin) that covalently links HA strands. TSG-6 has been used experimentally to treat sepsis and other forms of inflammation but its value in autoimmune diabetes is unknown. Finally, it may be possible to use HMW-HA to build immune tolerance to auto-antigens. We recently discovered that memory T-cells become IL-10-producing TR1 regulatory T-cells when they encounter their cognate antigen in the context of HMW-HA. Building on this finding, we have developed HMW-HA for use as a tolerizing adjuvant in an intranasal vaccine. Here, we will develop tolerizing vaccination as a tool for autoimmunity prevention. Our applications has three aims that each interrogate different aspects of the relationship between HMW-HA and regulatory T-cell function. They are: 1) to determine whether Treg in TID have impaired responses to HMW-HA, 2) to determine if strategies to promote HMW-HA integrity can prevent autoimmune diabetes, and 3) to develop a vaccine to promote auto-antigen specific immune tolerance using HMW-HA as a tolerizing adjuvant. The unifying goal of these studies is to use HMW-HA mediated tissue integrity signals to prevent autoimmune diabetes. PUBLIC HEALTH RELEVANCE: The proposed studies will greatly advance our understanding of the mechanisms that govern immune regulation in injured and healing pancreatic islets and will open the doors to innovative strategies to prevent autoimmune diabetes.